Effects of sampling strategy and DNA extraction on human skin microbiome investigations
Effects of sampling strategy and DNA extraction on human skin microbiome investigations
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DOI：
10.1038/s41598-019-53599-z
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发表时间：
2019-11-21
影响因子：
4.6
通讯作者：
Engstrand, Lars
中科院分区：
文献类型：
--
作者：
Bjerre, Rie Dybboe;Hugerth, Luisa Warchavchik;Engstrand, Lars
人类皮肤被多种微生物侵占，这些微生物在皮肤疾病中扮演着重要角色。研究皮肤微生物组提供了独特的障碍，如微生物生物量低。本研究的目的是建立皮肤微生物组分析的方法学，重点是采样技术和DNA提取。采集9名健康成人的皮肤拭子和刮痕，用12种商品试剂盒提取DNA。所有165个样本都使用16S rRNA基因进行了测序。比较eSwabs和SCRAPES捕获的种群，99.3%的序列是重叠的。使用eSwabs可以产生更高的一致性。用不同的DNA提取试剂盒制备文库的成功率为39%~100%。一些试剂盒具有较高的Shannonα多样性。对样本的一个子集(N=12)进行了超基因组分析。这些数据表明，在不降低16S rRNA文库制备成功率和不引入分类学偏见的情况下，将人类DNA从90%减少到57%是可行的。用拭子拭子是研究皮肤微生物群的可靠技术。DNA提取方法是成功测序的关键，并在微生物组分析中增加了大量的变异。应用元基因组学进行干预性研究时，建议减少宿主DNA。
The human skin is colonized by a wide array of microorganisms playing a role in skin disorders. Studying the skin microbiome provides unique obstacles such as low microbial biomass. The objective of this study was to establish methodology for skin microbiome analyses, focusing on sampling technique and DNA extraction. Skin swabs and scrapes were collected from 9 healthy adult subjects, and DNA extracted using 12 commercial kits. All 165 samples were sequenced using the 16S rRNA gene. Comparing the populations captured by eSwabs and scrapes, 99.3% of sequences overlapped. Using eSwabs yielded higher consistency. The success rate of library preparation applying different DNA extraction kits ranged from 39% to 100%. Some kits had higher Shannon alpha-diversity. Metagenomic shotgun analyses were performed on a subset of samples (N = 12). These data indicate that a reduction of human DNA from 90% to 57% is feasible without lowering the success of 16S rRNA library preparation and without introducing taxonomic bias. Using swabs is a reliable technique to investigate the skin microbiome. DNA extraction methodology is crucial for success of sequencing and adds a substantial amount of variation in microbiome analyses. Reduction of host DNA is recommended for interventional studies applying metagenomics.